The mechanisms through which a persistent recognition of oral commensal bacteria by oral epithelial cells (OECs) mitigates an uncontrolled inflammatory response of the oral mucosa remain unknown. Evidence suggests that epithelial cells may discriminate between commensals and pathogenic bacteria through differential activation of responses, particularly cytokine/chemokine patterns, which ultimately influence both innate and adaptive arms of the immune response. Recent work in our laboratory has been identifying patterns of OECs responses following challenge with commensal and pathogenic oral bacteria. The results showed a significantly more robust (quality and quantity) chemokine transcriptional activity in response to the oral commensal S. gordonii (Sg) compared to the outcomes following challenge with the oral pathogens F. nucleatum (Fn) and P. gingivalis. Subsequent protein analysis of selected chemokines showed that there was a clear disconnect between the Sg-induced chemokine transcriptional activation and the reduced and limited protein levels found in OEC extracts and supernatants in comparison with the effect of Fn, which despite inducing lower mRNA chemokines levels (2 to 4-fold less), enhanced a robust protein chemokine production as previously shown. Therefore, we hypothesized that Sg and perhaps other oral commensal bacterial species have the ability to efficiently activate regulatory mechanisms for chemokine transcription and translation in order to minimize pathological inflammatory responses driven by OECs in mucosal tissues. Consistently, preliminary analysis showed that Sg has the ability to significantly up-regulate (>2- fold) the expression of 115/2578 miRNAs in OECs, whereby miR-663a, miR-4516, miR492, and miR193a-5p have validated gene targets involved in TLR- and cytokine-induced chemokine transcription and translation. To test this hypothesis we propose the following two specific aims: (i) To determine the chemokine transcriptional and translational responses of OECs to oral commensal bacteria, and (ii) To determine the role of specific miRNAs in regulating the chemokine production induced by oral commensal bacteria in OECs. To address these knowledge gaps, we will use OEC cultures to: (a) determine whether the disconnect between transcription and translation of chemokines in OECs is common feature of oral commensal bacterial species, and (b) determine the effect of oral commensals on the expression of specific miRNAs as regulators of these inflammatory molecules. The contribution of this investigation is expected to be the identification of the activ regulation of inflammatory responses in OECs by specific miRNAs as a potential tolerogenic mechanism by which oral commensal bacteria maintain symbiosis with the host. Importantly, diminutions of miRNAs that would be activated specifically by oral commensal bacteria (e.g., during dysbiosis) could be related to pathologic inflammatory changes observed in disease. This project's significance will enable a better understanding of the cellular and molecular mechanisms involved in the tipping point between epithelial tolerogenic and pathologic responses to oral bacteria. The results are expected to contribute to a strong evidence-based foundation for future studies designed to target these mechanisms and identify new molecular target(s) associated with oral bacteria-regulated epithelial chemokine production, and will ultimately provide new opportunities for the development of innovative approaches to prevent/treat periodontitis.